Conventional HIV vaccine approaches have not achieved a desirable measure of protection in human clinical trials but certain inferences can be made. Stimulation of favorable innate immune responses will be needed to maximize the breadth and quality of adaptive immune responses. Long-term stimulation with antigen will be required to maintain a threshold level of protective immunity. Salient findings from our laboratories and other recent reports unify around these proposed requirements for a successful HIV vaccine. The genome of rhesus cytomegalovirus (RhCMV), the endogenous cytomegalovirus of rhesus macaques, has been engineered to express non-viral DNA sequences at high levels. Immunization with parental wild type (wt) RhCMV vectors expressing simian immunodeficiency virus (SIV) antigens induced resistance to infection after challenge with pathogenic SIVmac239 in a proportion (33%) of CMV-immunized animals (Hansen et al., 2009) possibly as a result of an antiviral T cell response that was skewed predominantly towards induction of SIV-specific effector memory T (Tem) cell responses. Deletion of the RhCMV-encoded viral interleukin (IL)-10 gene generated a potential vaccine vector (RhCMVdeltaIL10) with an attenuated replication phenotype that is still capable of inducing improved RhCMV-specific T cell responses, compared to parental RhCMV virus (Preliminary Data). Taken together, these findings provide a basis for further exploration and development of CMV-based vectors that are altered to improve safety and immunogenicity, and support the following hypothesis. A CMV vaccine vector based on an attenuated viral IL-10-deleted RhCMVdeltaIL10 vaccine vector that specifically activates innate immune mechanisms and improved T cell responses will induce enhanced antigen-specific immunity including amplified TEM cell responses. Accordingly, the improved immunogenicity imparted by a deletion of the viral IL-10 gene will generate a superior CMV-based vaccine vector for development of future HIV-1 vaccines. To address this hypothesis, rhesus macaques will be experimentally infected with either wild type parental RhCMV68-1 or attenuated RhCMVdeltaIL10 vaccine vectors and rigorously examined for antigen (RhCMV)-specific T cell responses in blood, lymph node, and rectal mucosa. RhCMVdeltaIL10 vector infection will be tested for persistence and immune activation in different tissue sites within the host (Aim 1). Recombinant RhCMVdeltaIL10 and RhCMV68-1 vectors expressing SIV antigens (Gag, Rev, Tat, Nef, or Env) will be constructed and analyzed in cell culture for expression of the SIV antigens, replication, and stability (Aim 2). These proposed studies will provide a preliminary investigation of immunogenicity for a new viral vector system and set the stage for future studies testing this vector modified for SIV antigens in rhesus macaques. Generation of such novel vectors carry the potential to significantly impact HIV vaccine design. PUBLIC HEALTH RELEVANCE: Studies proposed in this application will provide a preliminary investigation of a novel HIV-1 vaccine approach that incorporates a new viral vaccine vector, attenuated RhCMVdeltaIL10. Importantly, these studies will examine and develop a novel vaccine approach that is not previously tested in the SIVmac vaccine model, and carries the potential to significantly impact HIV vaccine design.